Physical properties of the plasma membrane appear to directly regulate the rates of several membrane mechanochemical functions; endocytosis, motility, and membrane resealing. In particular, these functions are inversely dependent upon the energy required to move an area of plasma membrane into a vesicle, lamellipodium or wound site. Using the laser tweezers, we can measure the force required to move plasma membrane into small membrane tubes or tethers. Indeed, we have shown that tether force correlates inversely with the rates of endocytosis, actin-dependent membrane extension and membrane resealing. Two factors contribute to the tether force: tension in the plasma membrane and membrane-cytoskeleton adhesion. Here we propose to measure the response of the whole cell to an increase in plasma membrane tension and to quantitatively analyze perturbations of membrane-cytoskeleton adhesion to develop a molecular model of adhesion. To test if membrane tension alone can inhibit endocytosis and motility, we will use high static force on tethers to create tension in the plasma membrane without swelling. It is believed that tension is continuous over the whole cell, and we will measure the coupling of oscillations in tension across the cell to test this. The degree of coupling will enable us to calculate the elastic parameters of spread cells from the known elasticity of the membrane. The energy of membrane-cytoskeleton adhesion appears to be set primarily by the level of phosphorylated inositol lipids, primarily phosphatidylinositol 4,5 diphosphate (PIP2). To test whether the free concentration of PIP2 or the surface charge of the plasma membrane correlates most closely with adhesion, we will measure adhesion as a function of cytoplasmic concentration of expressed EGFP-MARCKS effector or several different EGFP-PH domains. Microinjection of the same domains will enable us to determine if the time course of change in adhesion correlates best with the alteration of the cytoskeleton or direct inhibition of binding. Further, if adhesion recovers after microinjection, we will focus on the mechanism of recovery. These studies will provide the basis for a molecular model of adhesion, e.g. PIP2-PH domain bond density or actin filament density is the primary determinant of adhesion. Future studies will be focussed on refining the model of adhesion. Membrane tension, membrane-cytoskeleton adhesion and PIP2 levels are implicated in many cellular functions in addition to endocytosis, motility, and membrane resealing, e.g. volume regulation and exocytosis. Using quantitative physical and biochemical analyses of membrane parameters and cell activities, it will be possible to combine an understanding of the physical and biochemical activities to develop an integrated model of a given cell function.